Co-exposure to PS-NPs and HFPO-TA potentiates reproductive toxicity via ferroptosis and SLC1A5-Mediated glutamine deprivation in male mice

The pervasive co-contamination of ecosystems by nanoplastics (NPs) and per- and polyfluoroalkyl substances causes a synergistic harm to environmental and human health. In this study, murine and TM3 Leydig cell models were employed to explore the combined toxicities of polystyrene (PS)-NPs and hexafluoropropylene oxide trimer acid (HFPO-TA) to male reproduction and the potential mechanisms. Our results revealed that PS-NPs significantly exacerbated HFPO-TA-induced testicular histopathological damage, testosterone synthesis decline and spermatogenesis impairment. Mechanistically, HFPO-TA exposure triggered iron dyshomeostasis, mitochondrial dysfunction, oxidative stress and lipid peroxidation, culminating in Ferroptosis evidenced by Glutathione Peroxidase 4 (GPX4) downregulation and acyl-CoA synthetase long chain family member 4 (ACSL4) upregulation. Additionally, HFPO-TA directly bound to the glutamine transporter solute carrier family 1 member 5 (SLC1A5), suppressing glutamine uptake and glutathione biosynthesis. Crucially, co-treatment with PS-NPs synergistically amplified these alterations induced by HFPO-TA. Notably, both Ferroptosis inhibitor ferrostatin-1 and glutathione supplementation markedly attenuated the toxicity caused by PS-NPs and HFPO-TA co-exposure in Leydig cells, confirming Ferroptosis as the pivotal toxicological mechanism. Overall, PS-NPs aggravated Ferroptosis, glutathione deprivation and mitochondrial impairment upon HFPO-TA co-exposure in mouse testes and Leydig cells, indicating their synergy in inducing male reproductive toxicity. Our findings contribute to a deeper understanding regarding reproductive health risks from environmental co-exposure to NPs and HFPO-TA, and highlight Ferroptosis as a critical target pathway for intervention.